Various HPLC modes for the analysis of mono- and oligosaccharides.
•Precolumn derivatization with three novel UV absorbing tags was introduced.
•Sugars with charged tag were more readily separated by ion pair chromatography, IPC.
•The separation of negatively charged sugars by IPC was far superior than with WAX.
•Sugars with neutral tags resolved nicely using linear gradient of ACN on C18 column.
AbstractThree LC-based methods, including reversed-phase chromatography (RPC), ion-pair RPC and weak anion-exchange chromatography (WAX), were examined in the separations of precolumn derivatized mono- and oligosaccharides with the following three tagging agents: 1-naphthylamine (1-NA), 2-aminoanthracene (2-AA), and 3-amino-2,7-naphthalenedisulfonic acid (ANDSA). Due to differences in their charges and polarity, the three tagging agents imparted the sugar derivatives varying elution patterns in the three, just mentioned, chromatographic modes. While RPC yielded high resolution separations for 1-NA- and 2-AA-sugar derivatives, ion-pair RPC in the presence of the ion-pairing agent dodecyl trimethylammonium bromide (DTAB) in the mobile phase exhibited far more resolution and selectivity than WAX in the separation of ANDSA-sugar derivatives. This finding portrays the fact that an octadecyl column operating in ion-pair RPC mode can eliminate in most cases the need for an ion-exchange column for bioanalytical separations of ionic or ionizable species. Lastly, the characteristics of each chromatographic mode in the analysis of derivatized sugars are described using various mobile phase compositions.
KeywordsDerivatized sugars
UV-absorbing tags
Reversed-phase chromatography
Ion-pair chromatography
Anion-exchange chromatography
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